This invention relates generally to liquid chromatography and has particular reference to a novel method for indirect detection of nonelectrolytes.
The detection of nonelectrolytes is frequently limited by the insensitivity of the detector to the analyte. The problem is particularly acute for analytes which have no readily detectable spectroscopic, electrochemical or other property. The applicant is not aware of any prior art that has addressed itself to the problem of indirect detection of nonelectrolytes through the use of saturated additives in the eluent. The closest prior art known to the applicant is U.S. Pat. No. 3,929,410 issued to Schloss. This patent is directed to a method of indirect analysis where a saturated solution containing a known amount of radioactive standard is allowed to equilibrate with an unknown amount of the same compound in its unlabelled form, and quantitation of the unlabelled material is made on the basis of the amount of labelled material displaced. The present invention differs from that of the Schloss patent in that it does not require temperature changes to promote displacement of one material by another, is not restricted to radioactive materials, does not require the analyte and the standard to be of the same chemical structure, as will be described hereinafter.
The application of indirect detection to ion chromatography has been disclosed by Small and Miller in Canadian Pat. No. 1168705, and has been described by Cortes and Stevens (J. Chromatogr. 295(1984)269-275). In these references, a detectable ion is added to the eluent in a liquid chromatographic system, to serve as an indicator. When an analyte ion that is transparent to the detector is injected, it displaces an equivalent quantity of said indicator ion from the stationary phase. When the analyte enters the detector, the detector senses the decreased concentration of said indicator, and an indirect signal for the analyte ion is registered. This method is based on the principle of electroneutrality which requires the sum of the concentrations of analyte ion and indicator ion to be constant, since the concentration of the counterion, which is not retained by the stationary phase, is constant. The method applies only to ions, and cannot, therefore, be used for the detection of nonelectrolytes. As such, it differs from the present invention which is concerned only with the detection of nonelectrolytes.
The practice of indirect detection of nonelectrolytes has been described by Parkin (J. Chromatogr. 303(1984)436-439; J. Chromatogr. 287(1984)457-461), Parkin and Lau (J. Chromatogr. 314(1984)488-494), and Vigh and Leitold (J. Chromatogr. 312(1984)345-356). In these references, a detectable nonelectrolyte, hereinafter referred to as the additive, is added to the eluent to serve as an indicator. When a transparent analyte is injected, it disturbs the partitioning of the additive between the stationary phase and the eluent. When the analyte enters the detector, the detector senses the altered concentration of the additive in the eluent, and generates a signal. The sensitivity of the procedure is poor, as explicitly recognized by Parkin (J. Chromatogr. 303(1984)436-439). The present invention differs from this prior art method in the important regard that the additives used are present at saturation in the eluent. This novel feature leads to a very substantial improvement in the sensitivity of detection as will be described hereinafter.
Other prior art developed in the course of a preliminary search consists of U.S. Pat. Nos. 4,223,020; 3,222,525; 2,954,333 and 3,623,840.